Cullins (CULs) are members of an evolutionary conserved family of proteins that form the core of multisubunit ubiquitin ligases. CUL1 and CUL3, associate with a large number of different adapters that appear to target specific substrates for degradation. However, only few substrates of cullin ubiquitin ligases are currently known. Equally unclear are the mechanisms that regulate the assembly of cullins into substrate specific ubiquitin ligase complexes, a process which is thought to require the controlled exchange of labile adapters. Recent evidence suggests that the COP9 signalosome (CSN) may participate in this process by keeping cullins in an inactive state. The goal of these studies is to identify mechanisms regulating cullin ubiquitin ligase assembly and substrate-specific function in fission yeast. We are testing the hypothesis that enzymatic activities associated with CSN neutralize spurious cullin activity, in order to provide a safe environment for CUL1 and CUL3 ubiquitin ligase assembly by preventing autocatalytic destruction of adapter proteins. In addition, we are pursuing the hypothesis that CUL4 also assembles in a similarly controlled manner into multisubunit ligases that target distinct substrates that remain to be identified. The experimental approach includes (1.) Testing the role of CSN in the assembly and function of CUL1 and CUL3 complexes, (2.) identifying and characterizing the components of CUL4-associated ubiquitin ligases, and (3.) identifying and characterizing CUL3 and CUL4 substrates.